![Fig. 3. Activation of HPg by Sak. Human wtr-E1-HPg (10 nmol/L) was activated by Sak (10 nmol/L) in the presence of the chromogenic substrate, S2251 (0.9 mmol/L). The HPm generated as a function of time was detected by the liberation of p-nitroanilide from the substrate and is depicted here as the absorbance at 405 nm. The curves illustrated are for wtr-E1-HPg, wtr-E1-HPg to which a small amount (0.5 nmol/L) of wtr-HPm was added at the 6-minute time point of the incubation of Sak and wtr-E1-HPg, and for the mutants, r-[R561A]E1-HPg and r-[D646E]E1-HPg. The reduced DodSO4/PAGE gel insert shows the protein components present after 30 minutes of activation. The buffer was 10 mmol/L HEPES-NaOH, pH 7.4, at 37°C. The activation of wtr-E1-HPg by lmw-uPA displayed in the gel insert was accomplished as in Fig 1.](https://ash.silverchair-cdn.com/ash/content_public/journal/blood/89/5/10.1182_blood.v89.5.1585/3/bl_0001f3.jpeg?Expires=1724288841&Signature=j71Bjy~kaabdhFYZhgmXZpgBmByOcPV3nVOj~imLIWIo8RR-KkT2qn1uQCJ-cHOk-RMyxwnVzATENVgr1DIlY4zGhtVB-NW6sMqxwu8FDedRfVZBpKG6YdpUhap4ZgPW8c0~KgVgMN-KA3SorlJRxizFsi-NejLCV0BAZHMhQCpNCTBoVNYuV41FyjmkTsVKhEfWbARo0rMQB2mCrnwrIqO0Q3H-i2tNOp8wWFbRi02IFAlDLoLPQ3OdjavukeSHESaakiwveZS6BKuPxN~K9eia23fZecJ-kcEKKsoVx1FYlUyvOkzYzrl0gkIjFDwuWwmB7MB34UK-qBae6jJcEw__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Activation of HPg by Sak. Human wtr-E1-HPg (10 nmol/L) was activated by Sak (10 nmol/L) in the presence of the chromogenic substrate, S2251 (0.9 mmol/L). The HPm generated as a function of time was detected by the liberation of p-nitroanilide from the substrate and is depicted here as the absorbance at 405 nm. The curves illustrated are for wtr-E1-HPg, wtr-E1-HPg to which a small amount (0.5 nmol/L) of wtr-HPm was added at the 6-minute time point of the incubation of Sak and wtr-E1-HPg, and for the mutants, r-[R561A]E1-HPg and r-[D646E]E1-HPg. The reduced DodSO4/PAGE gel insert shows the protein components present after 30 minutes of activation. The buffer was 10 mmol/L HEPES-NaOH, pH 7.4, at 37°C. The activation of wtr-E1-HPg by lmw-uPA displayed in the gel insert was accomplished as in Fig 1.
